High-performance Thin-layer Chromatography Technique Research Articles

Simultaneous Estimation of Monoterpenes (Eugenol, Eucalyptol and R-Limonene) by HPTLC Densitometric Method and Its Validation.

Plant-derived monoterpenes have various pharmacological applications, but the problem lies with their detection and estimation. Hence, it is crucial to develop a robust technique to overcome this problem. This work was aimed at developing a swift, accurate and reproducible technique for the simultaneous detection of three monoterpenes, namely, eugenol, eucalyptol (1,8- cineole), and R-limonene, using the high-performance thin-layer chromatography technique, as per International Council of Harmonization (ICH) guidelines. The best results were obtained in the eluent mixture prepared by using hexane: toluene: ethyl acetate (6:3:1, v/v/v), and silica gel-coated aluminum thin layer chromatography (TLC) plates 60F254 as a stationary phase that produced very sharp and well-resolved symmetrical peaks for eugenol, eucalyptol, and R-limonene at Rf values of 0.47 ± 0.03, 0.56 ± 0.03, and 0.71 ± 0.03, respectively. The linear range for eugenol, eucalyptol, and R-limonene was 1 to 15 ng/ spot (r2 = 0.9906), 100 to 600 ng/spot (r2 = 0.99625), 50 to 300 ng/spot (r2 = 0.9873), respectively. Furthermore, the limit of detection (LoD) and limit of quantitation (LoQ) values for eugenol were obtained at 0.0129 and 0.0391 ng/spot, followed by eucalyptol (0.82 and 2.48 ng/spot) and R-limonene (0.594 and 1.8 ng/spot), respectively. To our knowledge, we are the first to report a quick and reproducible high-performance thin-layer chromatography (HPTLC) technique for the simultaneous determination of eugenol, eucalyptol (1,8-cineole), and R-limonene. This method can further be used for the quantification of these secondary metabolites in plant extracts, enriched fractions, and pharmaceutical formulations.

Simultaneous Estimation of Monoterpenes (Eugenol, Eucalyptol And R-Limonene) by HPTLC Densitometric Method and Its Validation .

Plant-derived monoterpenes have various pharmacological applications, but the problem lies with their detection and estimation. Hence, it is crucial to develop a robust technique to overcome this problem. This work was aimed at developing a swift, accurate and reproducible technique for the simultaneous detection of three monoterpenes, namely, eugenol, eucalyptol (1,8-cineole), and R-limonene, using the high-performance thin-layer chromatography technique, as per ICH guidelines. The best results were obtained in the eluent mixture prepared by using hexane: toluene: ethyl acetate (6:3:1, v/v/v), and silica gelcoated aluminum TLC plates 60F254 as a stationary phase that produced very sharp and well-resolved symmetrical peaks for eugenol, eucalyptol, and R-limonene at Rf values of 0.47 ± 0.03, 0.56 ± 0.03, and 0.71 ± 0.03, respectively. The linear range for eugenol, eucalyptol, and R-limonene was 1 to 15 ng/spot (r2 = 0.9906), 100 to 600 ng/spot (r2 = 0.99625), 50 to 300 ng/ spot (r2 = 0.9873), respectively. Furthermore, the LoD and LoQ values for eugenol were obtained at 0.0129 and 0.0391 ng/spot, followed by eucalyptol (0.82 and 2.48 ng/spot) and R-limonene (0.594 and 1.8 ng/spot), respectively. To the best knowledge, we are the first to report a quick and reproducible HPTLC technique for the simultaneous determination of eugenol, eucalyptol (1,8-cineole), and R-limonene. This method can further be used for the quantification of these secondary metabolites in plant extracts, enriched fractions, and pharmaceutical formulations.

Impact of altitudinal variation on secondary metabolites, antioxidant, anti-inflammatory and anticancer potential of Adiantum venustum D. Don

Phytoconstituents are being emphasized as chemotherapeutic agents, as well as probable anticancer agents. The purpose of the present research was to assess the antioxidant, anti-inflammatory, and anticancer properties of crude ethanolic extracts and bioactive fractions of Adiantum venustum D. Don collected from three sites in Himachal Pradesh, India. The bioactive phytoconstituents were examined qualitatively and quantitatively in crude ethanolic extracts and fractions of all sites, and biological activities were analyzed. The potential presence of terpenoids, saponins, tannins, phenols, and flavonoids was examined, and it came to light that BF-II [n-butanol fraction of Site-II (2000 m altitude)] had a significantly higher phytochemical concentration. Additionally, the crude ethanolic extracts and fractions were examined for antioxidant and anti-inflammatory properties, and BF-II was found to have potential antioxidant and anti-inflammatory activity, as verified by the IC50 values (lowest half inhibitory concentration) in the DPPH (57.86 ± 0.72 μg/mL), reducing power assay (45.20 ± 0.72 μg/mL), FRAP (73.12 ± 0.23 μM Fe equivalent), BSA (56.76 ± 0.25 μg/mL), and EAA (EAA59.84 ± 0.13 μg/mL). Following that, the anticancer potential was investigated further using the MTT assay on the HeLa cell line, MTT assay revealed that BF-II possesses significantly higher anticancer potential. Furthermore, the dominant crude ethanolic extract and fraction were further investigated using gas chromatography-mass spectroscopy, and high-performance thin layer chromatography techniques from the selected plant. Overall, the study suggests that the n-butanol fraction of A. venustum possesses significantly higher therapeutic value but is still not explored to the extent of its potential.

Phytochemical screening and pharmacognostical evaluation of cordia sebestena Linn.

Plants are known as treasures of effective drugs for various ailments from ancient times. Plant metabolites played a protagonist part in the detection and development of novel chemotherapeutic mediators. Therefore, exploring the medicinal importance of plants has become extremely important concerning the evolving ailments of the present times. In the existing investigation, phyto-constituents and antioxidant activity of the plant known as Cordia sebestena Linn. were discovered for their biological importance. Cordia sebestena Linn. is an understudied medicinal plant that belongs to the family Boraginaceae and is commonly known as Aloe wood, geranium tree, and sea trumpet. The investigation objective is to develop the high-performance thin layer chromatography (HPTLC) fingerprint outline of the phytoconstituent of Cordia sebestena Linn, to identify the component of medicinal importance. In the present study, the HPTLC technique for the parting of the active constituents in extracts has been developed via the solvent system toluene: methanol: acetone (16:4:1). This study has provided the biological source which can be an eco-friendly, cost-effective, and better option than the chemically synthesized drugs. In the future, these fingerprinting descriptions will be supportive in further studies like quality regulation of the drug and estimating therapeutic worth.

Simultaneous determination of lesinurad and febuxostat in commercial fixed-dose combinations using a greener normal-phase HPTLC method

Abstract So far, no documented method for simultaneously analyzing lesinurad (LND) and febuxostat (FBX) has been reported for either traditional high-performance thin-layer chromatography (HPTLC) or a green HPTLC technique. In order to determine LND and FBX simultaneously in commercially available fixed-dose combo tablets, this study devised a normal-phase HPTLC method that is fast, sensitive, and green. The green eluents for the simultaneous analysis of LND and FBX were a mixture of ethyl acetate:ethanol:water at 70:20:10 (v/v/v) ratio. The new approach’s greenness was predicted utilizing four distinct greenness tools: the National Environmental Method Index, Analytical Eco-Scale, ChlorTox, and Analytical GREENness approaches, and the results revealed a significantly greener profile. The current method operated on a linear scale between 30 and 1,000 ng·band−1. It was confirmed that the current approach is sensitive, accurate, precise, robust, and green. The LND and FBX contents of commercially available tablet products A and B were found to be within the range of 100 ± 2%, indicating that the existing methodology for simultaneously determining LND and FBX in pharmaceutical combination products is applicable. The results of the current methodology indicated that LND and FBX could be consistently measured in pharmaceutical combination products simultaneously using the current approach.

Development of Green HPTLC method for simultaneous determination of a promising combination Tamsulosin and Mirabegron: stability-indicating assay was examined

Recently, mirabegron has been added to tamsulosin to treat overactive bladder in men with benign prostatic hypertrophy. A Rapid, selective, sensitive, and green high-performance thin-layer chromatography (HPTLC) approach was developed for the simultaneous determination of tamsulosin (TAM) and mirabegron (MIR) in pure and laboratory-prepared mixture. Complete separation was obtained on silica gel F254 using the solvent system methanol-ethyl acetate-ammonia (3:7:0.1, v/v). Short-wave ultraviolet light at 270 nm was used to view the chromatographic bands. For MIR and TAM, the suggested technique revealed compact spots with retention factor Rf values of 0.42 and 0.63, respectively. Within concentration ranges of 0.15–7.5 µg/band and 0.05–2.5 µg/band, good linearity was observed, with mean percentage recoveries of 100.04 ± 0.56 and 99.98% ± 0.95 for MIR and TAM, respectively. Green assessment of the developed HPTLC technique was estimated using different green analytical chemistry metrics such as Analytical eco-scale Analytical GREEness (AGREE), and Green Analytical Procedure Index (GAPI) metrics. The proposed method was effectively used as a stability-indicating assay to assess the presence of MIR and TAM in the pharmaceutical dosage form in the presence of their degradation product. The statistical analysis showed high precision and accuracy.

A high performance thin layer chromatography (HPTLC) method for the quality assessment of citronella oil: application in commercial sample analysis

Citronella oil, extracted from Cymbopogon species (winterianus and nardus) is a commercially valuable essential oil used in personal-care products and insect repellents. Routine analysis in gas chromatography is incapable of detecting high-boiling adulterants therein. In this study, an HPTLC technique was developed for the absolute quantification of citronellal (characteristic chemical marker) and triglyceride (main constituent of vegetable oil adulterant) in citronella oil for its quality assessment. It was validated in terms of specificity, linearity, sensitivity, accuracy and precision. Further, the developed method was employed to quantify citronellal and triglyceride in twenty commercial samples. The results showed a wide variation in citronellal content (trace to 30.65% w/w) and could differentiate its two chemotypes. Also, it revealed the possibility of vegetable oil adulteration through the detection and quantification of triglyceride in selected samples. It can be a simple and rapid technique for the quality control of citronella oil.

High-Performance Thin-Layer Chromatography hyphenated with image processing and chemometrics as a tool for forensic discrimination of Cannabis sativa L. chemotypes

Cannabis sativa L. is the most widely cultivated, trafficked, and abused illicit drug. The reliable distinction between drug type and fiber type C. sativa remains a topic of interest for forensic chemists. For the first time, we applied a combination of simple, cost-effective, and reliable High-Performance Thin-Layer Chromatography (HPTLC) with image analysis as well as sophisticated multivariate tools for differentiating C. sativa drug, fiber and intermediate chemotypes. After extraction, major cannabinoids from 43 seized C. sativa were separated using the HPTLC method. Peak areas of Δ9-THC, CBN, and CBD were calculated by simple image processing, and Xfactor = [THC + CBN]/CBD was determined as criteria used to discriminate three C. sativa chemotypes. The obtained results were compared with Gas Chromatography with Flame Ionization Detection (GC-FID) as the reference method for the determination of Xfactor.Principal Component Analysis and Hierarchic Cluster Analysis were applied to classify three C. sativa chemotypes according to their chemical pattern. The proposed approach clearly distinguishes 26 drugs, 13 fiber, and 4 intermediate chemotypes in seized C. sativa, aligning with GC-FID analysis. The results showed that the HPTLC technique in combination with multivariate methods is an accurate and reliable tool for high-throughput and forensic screening of three C. sativa chemotypes. Compounds such as Δ9-THC and CBD were marked as the most important cannabinoids responsible for the classification of the seized C. sativa chemotypes.

A Review of various Analytical Methods Developed for Estimation of Paracetamol, Diclofenac sodium, and Chlorzoxazone in Single-component and Multi-component Dosage Form

Painful muco-skeletal joint disorders such as osteoarthritis, rheumatoid arthritis, and ankylosing spondylitis are typically treated with a combination of paracetamol, diclofenac sodium, and chlorzoxazone. Paracetamol and diclofenac sodium are non-steroidal anti-inflammatory drugs, while chlorzoxazone is a muscle relaxant. It acts on the central nervous system and the spinal cord to reduce muscle stiffness and spasms as well as increase muscle mobility. The most commonly used pharmaceutical medicine is paracetamol, which comes in a variety of dosage forms such as injection, tablet, capsule, drops, elixirs, suspensions, and suppositories. However, due to toxicity, it may cause mortality in large dosages with extended intake. As a result, more efficient analytical approaches aimed at providing quality control of routinely used medications are required. Numerous analytical methods, both spectroscopic and chromatographic such as high-performance liquid chromatography, gas chromatography with mass spectrometry, and high-performance thin-layer chromatography have been established to estimate paracetamol diclofenac sodium and chlorzoxazone in their single-component dosage form. The simultaneous estimation of paracetamol, diclofenac sodium, and chlorzoxazone in combined dosage forms has been performed using reverse-phase high-performance liquid chromatography and high-performance thin-layer chromatography techniques. In contrast, there are very few techniques available to estimate paracetamol, diclofenac sodium, and chlorzoxazone in combination pharmaceutical dose form till date. This article is an attempt to jut out all the methods developed to estimate the above-mentioned drugs in the single or multi-component dosage form.

Separation and quantification of azelnidipine and chlorthalidone in a synthetic mixture using optimized HPTLC method

A simple, accurate, and reliable high-performance thin-layer chromatographic technique has been developed and validated for the simultaneous quantitation of azelnidipine and chlorthalidone in bulk and synthetic mixtures. The procedure was carried out using a precoated silica gel 60 F254 TLC plate with a mobile phase of chloroform, ethyl acetate, and methanol in the ratio of 6.5:3.5:0.6 (by volume). Thin-layer chromatographic densitometry at 240nm was used to quantify medicines chromatographically. Over concentration ranges of 250.0 to 1000.0ng/band for chlorthalidone and 160.0 to 640.0ng/band for azelnidipine, the high-performance thin-layer chromatography technique was quantitated. This technique produced a tight and well-resolved band at retention factors of 0.67±0.02 and 0.24±0.02 for azelnidipine and chlorthalidone, respectively. Data from a linear regression study calibrating this method revealed a strong linear correlation between the two approaches, with regression coefficients of r2>0.99 for both. According to The International Conference for Harmonization of Technical Requirements for Pharmaceuticals for Human Use requirements, the procedures were validated for precision, robustness, accuracy, and specificity. The developed method was also used to simultaneously estimate azelnidipine and chlorthalidone in a synthetic mixture. The results were found to be in exemplary % assay with label claims.

Phytochemical study, antioxidant and vasodilatation activities of leafy stem extracts of Flemingia faginea Guill. & Perr. (Barker), a medicinal plant used for the traditional treatment of arterial hypertension

BackgroundFlemingia faginea Guill. & Perr. is a medicinal plant used for the endogenous treatment of hypertension. This study aimed to establish the phytochemical profile, antioxidant power, and vasodilation effects of aqueous decoction (FAD), ethyl acetate (EAF) and aqueous residual fractions (ARF) of the leafy stem of Flemingia faginea (F. faginea). MethodsThe phytochemical study was performed using high-performance thin-layer chromatography technique and high-performance liquid chromatography coupled with mass spectrometry (HPLC-MS). Folin-Ciocalteu and aluminum trichloride colorimetric methods were used, respectively, to quantify total polyphenols and flavonoids. The antioxidant properties of the extracts were determined using 2,29-azinobis-3-ethylbenzothiazoline-6-sulfonic (ABTS•), 2,2-diphenyl-1-picrylhydrazyl (DPPH•) and ferric ion reduction (FRAP) methods. The vasodilatory effects of F. faginea extracts were performed using the DMT 620M myograph technique for an ex vivo study. ResultsThe phytochemical study revealed the presence of flavonoids, tannins, coumarins, sterols, and triterpenes, alkaloids, and saponins in the extracts studied. HPLC MS analysis identified as the main phenolic compounds, gallic acid-glucoside (16.56 ± 0.06 mg/g), myricetin rhamnoside (15.39 ± 0.04 mg/g), myricetin rutinoside (9.32 ± 0.00 mg/g), quercetin rutinoside (9.05 ± 0.00 mg/g), caffeoyl-glucoside (6.07 ± 0.02 mg/g) and 5-Caffeoylquinic acid (4.52 ± 0.00 mg/g). The present data also showed higher contents of phenolic compounds and flavonoids in fractions of ARF (280.49 ± 0.08 mgGAE/g) and EAF (33.31 ± 0.33 mgQE/g) fractions respectively. Furthermore, the ARF fraction had the best iron ion reduction capacity (16.96 ± 0.01µgAAE/100 mg), while, the EAF (ABTS: IC50=67.33 ± 0.04 µg/mL; DPPH: IC50=4.69 ± 0.20 µg/mL) was the best free radical scavenger. In endothelium-intact rings, study extracts showed a vasodilation effect in a concentration-dependent manner with half-maximum effective concentration (EC50) of 0.24 ± 0.03 mg/mL, 0.85 ± 0.07 mg/mL and 0.05±0.01 mg/mL respectively for FAD, ARF and EAF. In endothelium-denuded vessels, these values were 0.26 ± 0.09 mg/mL, 0.57 ± 0.05 mg/mL, and 0.85 ± 0.01 mg/mL for EAF, FAD, and ARF, respectively. The maximum effects were of 99.94 ± 0.15%, 86.89 ± 5.58%, and 62.69 ± 5.14% (endothelium-intact) and of 100 ± 0.06%, 96.55 ± 4.68%, and 74.16 ± 4.06% (endothelium-denuded), respectively, for EAF, FAD, and ARF with a significant difference between them. ConclusionThis study reports that the decoction and fractions of F. faginea are rich in various chemical compounds and possess antioxidant and vasodilatory properties. These preliminary data provide a scientific basis which contributed to justify that plant used in traditional medicine.

Phenolic Profile and In Vitro Antioxidant Activity of Different Corn and Rice Varieties.

Celiac disease (CD) is an autoimmune disease. To date, the only universally recognized treatment for CD is the gluten-free diet (GFD). Despite the GFD, a state of inflammation and oxidative stress could remain at the intestinal level of celiac patients. Several components of the diet, such as phenolic compounds with known antioxidant properties, could play a protective role in the inflammatory state of patients with CD. The objective of this study was the characterization of the phenolic profile and the antioxidant capacity of pigmented cereals (rice and corn) from the Italian market and farms. Different in vitro methods were applied: Folin-Ciocalteu assay, pH differential method, DPPH assay, TEAC assay, and High-Performance Thin Layer Chromatography technique. According to the results, pigmented varieties are possible valuable sources of phenolic compounds and anthocyanins with high antioxidant activity. They could be used as alternative ingredients for the formulation of gluten-free products.

Determination of Cordycepin Using a Stability-Indicating Greener HPTLC Method

A wide range of analytical techniques have been reported to determine cordycepin (CDN) in various sample matrices. Nevertheless, greener analytical approaches for CDN estimation are scarce in the literature. As a result, this study was designed to develop and validate a stability-indicating greener “high-performance thin-layer chromatography (HPTLC)” technique for CDN determination in a laboratory-developed formulation. The greener eluent system for CDN detection was ethanol–water (75:25 v/v). At a wavelength of 262 nm, CDN was measured. The greenness scale of the proposed analytical technology was derived using the “Analytical GREENness (AGREE)” approach. The proposed stability-indicating HPTLC assay was linear for CDN analysis in the 50–1000 ng/band range with a determination coefficient of 0.9978. The proposed analytical technique for CDN analysis was simple, rapid, accurate, precise, robust, selective, stability-indicating, and greener. The AGREE score for the proposed stability-indicating HPTLC technique was calculated to be 0.79 using the AGREE calculator. The current protocol was able to detect CDN degradation products under various stress conditions, indicating its stability-indication characteristics and selectivity. The AGREE quantitative score indicated that the stability-indicating current protocol had outstanding greener characteristics. The amount of CDN in the laboratory-developed formulation was determined to be 98.84%, indicating the suitability of the current protocol in the assay of CDN in the formulations. These results suggested that CDN in a laboratory-developed formulation may be regularly determined using the stability-indicating greener HPTLC strategy.

Comparative phytochemical screening through high-performance thin layer chromatography technique and free radical scavenging ability of five species of genus Clerodendrum

Comparative phytochemical screening through high-performance thin layer chromatography technique and free radical scavenging ability of five species of genus Clerodendrum

Determination of Colchicine in Pharmaceutical Formulations, Traditional Extracts, and Ultrasonication-Based Extracts of Colchicum autumnale Pleniflorum (L.) Using Regular and Greener HPTLC Approaches: A Comparative Evaluation of Validation Parameters.

In the literature, there is a scarcity of greener analytical approaches for colchicine (CLH) analysis. As a result, efforts were made in this study to develop and validate a greener reversed-phase high-performance thin-layer chromatography (HPTLC) technique for CLH analysis in traditional extracts (TE) and ultrasonication-based extracts (UBE) of commercial Unani formulations, commercial allopathic formulations, and Colchicum autumnale Pleniflorum (L.) obtained from Egypt and India. This new technique was compared to the regular normal-phase HPTLC method. The greenness profile of both methods was estimated using the Analytical GREENness (AGREE) approach. In the 100–600 and 25–1200 ng/band ranges, regular and greener HPTLC procedures were linear for CLH analysis, respectively. For CLH analysis, the greener HPTLC method was more sensitive, accurate, precise, and robust than the regular HPTLC method. For CLH analysis in TE and UBE of commercial Unani formulations, commercial allopathic formulations, and C. autumnale obtained from Egypt and India, the greener HPTLC method was superior in terms of CLH content compared to the regular HPTLC method. In addition, the UBE procedure was superior to the TE procedure for both methods. The AGREE scores for regular and greener reversed-phase HPTLC methods were found to be 0.46 and 0.75, respectively. The AGREE results showed excellent greener profile of the greener HPTLC method over the regular HPTLC technique. Based on several validation criteria and pharmaceutical assay findings, the greener HPTLC method is regarded as superior to the regular HPTLC approach.

Quantification of biomarkers and evaluation of antioxidant, anti-inflammatory, and cytotoxicity properties of Dodonaea viscosa grown in Saudi Arabia using HPTLC technique

Abstract Dodonaea viscosa (Sapindaceae) was collected from Riyadh, Saudi Arabia. For the simultaneous measurement of quercetin and kaempferol, a validated high-performance thin-layer chromatography (HPTLC) approach was devised in D. viscosa leaf extract. The antioxidant activity was evaluated using diphenyl 1-picrylhydrazyl (DPPH) and 2,2′-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS) assays. Moreover, the cytotoxic effect was tested against three cancer cell lines A549, HepG2, and MDA-MB-231. The potential anti-inflammatory properties of different fractions of D. viscosa were also evaluated using lipopolysaccharide (LPS)-induced THP-1 macrophages cells. The test samples include a crude extract of leaves and its solvent-soluble fractions of D. viscosa. The results showed that the crude extract and its fractions exhibited various significant biological activities, the fraction of chloroform demonstrated the highest free radical scavenging activity with IC50 values: 172.2 and 257.7 µg/mL for both DPPH and ABTS tests. Additionally, the chloroform fraction had the greatest cytotoxic activity against MDA-MB-231 (IC50 values: 24.6 ± 0.4 µg/mL). Moreover, the chloroform fraction exhibited the highest downregulation of the LPS-induced expression of TNF-α and IL-6. Quercetin and kaempferol were estimated concurrently in leaves crude extract using a validated technique on an HPTLC plate (10 cm2 × 10 cm2) with a combination of toluene–ethyl acetate–formic acid (5:4:0.2; v/v/v) as the mobile phase and a λ max of 254 nm. The amount of quercetin and kaempferol was found to be 31.8 and 15.01 mg/g of dried leaf extract, respectively. The presence of high levels of quercetin and kaempferol in D. viscosa leaves extract could explain its remarkable antioxidant, cytotoxic, and anti-inflammatory effects. The devolved HPTLC method can be used for routine analysis and standardization of D. viscosa crude plant material, extracts, and/or finished products using quercetin and kaempferol as appropriate markers.

A Simple, Cost-Effective, and Green HPTLC Method for the Estimation of Ascorbic Acid in Solvent and Ultrasound-Assisted Extracts of Phyllanthus emblica, Capsicum annuum, and Psidium guajava

Greener analytical methodologies for the estimation of ascorbic acid (AA) are poorly reported in the literature. Furthermore, the green indexes of the literature’s analytical assays of AA estimation have not been assessed. As a consequence, the aim of this research is to invent and validate a simple, cost-effective, and green reverse-phase “high-performance thin-layer chromatography (HPTLC)” method for the estimating AA in the solvent extracts (SE) and ultrasound-assisted extracts (UAE) of Phyllanthus emblica, Psidium guajava, and Capsicum annuum. The greener mobile phase for AA estimation was a binary mixture of water and ethanol (70:30, v/v). At a wavelength of 265 nm, the detection of AA was carried out. The greener HPTLC technique was linear in the 25–1200 ng/band range. In addition, the method was simple, cost-effective, accurate, precise, robust, sensitive, and green. The amount of AA was highest in the SE and UAE of P. emblica compared to the SE and UAE of P. guajava and C. annuum. The amount of AA in the SE of P. emblica, P. guajava, and C. annuum was found to be 491.16, 168.91, and 144.30 mg/100 g, respectively. However, the amount of AA in the UAE of P. emblica, P. guajava, and C. annuum was found to be 673.02, 218.71, and 199.30 mg/100 g, respectively. Using the “analytical GREEnness (AGREE)” methodology, the greenness index for the developed method was calculated to be 0.88, showing that the developed method has an excellent green profile. When it came to extracting AA, the UAE method outperformed the SE method. These findings suggested that the developed method might be used to estimate the AA in a variety of vegetable crops, plant-based extracts, and commercial formulations. Furthermore, because of the use of greener solvent systems against the commonly utilized hazardous solvent systems for AA determination, this technique is also safe and sustainable.

Antidiabetic Potential of Ficus glomerata Roots with a Special Emphasis on Estimation of Bioactive Compounds by a Novel Validated HPTLC Technique

Abstract: Background: The data presented in this article for Ficus glomerata Linn. belonging to family Moraceace which is commonly found all over India. This study aimed towards the development and validation of high-performance thin-layer chromatography (HPTLC) method for simultaneous estimation of lupeol and quercetin from Ficus glomerata and correlate with its antidiabetic potential. Methods: The various fractions of ethanolic extract of Ficus glomerata root were prepared. The HPTLC analysis of quercetin and lupeol which are the important phytoconstituents responsible for various pharmacological actions was carried out at 525 nm. ICH guidelines were followed to validate this method for accuracy, precision and repeatability. Results: The linearity range of quercetin and lupeol were obtained as 400-2400 ng/spot and 1000- 5000 ng/spot respectively. Percent drug content was highest in diethyl ether fraction (quercetin 2531.8 ng and lupeol 1400 ng). The limit of detection value (LOD) obtained for quercetin and lupeol was 3.0793 and 3.1645 ng and the limit of quantification (LOQ) was 9.3314 and 9.5895 ng respectively. This method developed was accurate, precise and simple has shown higher resolution from other phytoconstituents present in the fractions. The method can be very effectively applied for analyzing the quality of herbal material and formulations containing Ficus glomerata. Antidiabetic activity of various fractions of ethanolic extract of Ficus glomerata roots was studied on alloxan-induced diabetic rats. Treatment with fractions was continued for 11 days. The effect of the fractions on glucose was analyzed. Diabetic rats treated with diethyl ether fraction exhibited a significant (p<0.05) decrease in glucose levels, indicating the potential use of Ficus glomerata in diabetes mellitus. Conclusion: As per the ICH guidelines, the HPTLC method used for simultaneous estimation of lupeol and quercetin was accurate, precise and specific. The method used for phytochemical standardization of various fractions of ethanolic extract of the roots of Ficus glomerata and correlated with its antidiabetic activity. Key words: Ficus glomerata, HPTLC, Lupeol, Quercetin, Alloxan, Antidiabetic activity

Quantitative Phytochemical Analysis and in vitro Study of Antioxidant and Anti-inflammatory Activities of Aegle marmelos Fruit with Peel and without Peel: A Comparative Evaluation

Background: Aegle marmelos is an important ethnomedicinal plant that is found commonly in India and other Asian countries. The peel of the fruit is hard and is often discarded as waste. However, a few studies have shown that even the peel has antioxidant and anti-inflammatory attributes. The aim of this research work was to perform a comparative evaluation of Aegle marmelos fruit extracts with the peel (whole fruit) and without the peel. Phytochemical investigations were carried out on the two extracts. Materials and Methods: The Folin-ciocalteu reagent method and the aluminum chloride colorimetric method were used to determine the total phenolic contents and total flavonoid contents, respectively. The marmelosin content was measured using the high-performance thinlayer chromatography technique. The antioxidant activity was determined in-vitro using the free-radical scavenging method, and the in vitro antiinflammatory property was determined using the egg albumin denaturation method. The unpaired t-test (with Welch’s correction) was used to determine P values. Any differences that were observed were considered significant if P &lt; 0.05. Results: The total flavonoid content of the extracts was found to be low, while the total phenolic content was high. There were no significant differences in the antioxidant activities of the extracts. The anti-inflammatory activities and marmelosin contents of the extracts were significantly different. Conclusion: The extracts displayed good antioxidant and anti-inflammatory activities. The anti-inflammatory activity of the extract of the fruit without the peel was found to be greater than that of the extract of the fruit with the peel.

Determination of Thymol in Commercial Formulation, Essential Oils, Traditional, and Ultrasound-Based Extracts of Thymus vulgaris and Origanum vulgare Using a Greener HPTLC Approach.

In the literature, greener analytical approaches for determining thymol in its commercial formulations, plant-based phytopharmaceuticals, and biological fluids are scarce. As a result, the goal of this study is to develop and validate a normal-phase “high-performance thin-layer chromatography (HPTLC)” method for determining thymol in commercial formulations, essential oils, traditional extracts (TE), and ultrasound-based extracts (UBE) of Thymus vulgaris and Origanum vulgare obtained from various geographical regions. The greener mobile phase for thymol analysis was a binary combination of cyclohexane and ethyl acetate (85:15, v/v). The derivatized densitometric analysis of thymol was carried out under visible mode at 530 nm utilizing anisaldehyde-sulfuric acid as a derivatizing/visualizing agent. In the 10–2000 ng/band range, the greener normal-phase HPTLC method was linear. Furthermore, for thymol analysis, the proposed analytical approach was simple, quick, inexpensive, accurate, precise, robust, sensitive, and greener. The thymol contents in commercial formulation were computed as 7.61% w/w. In general, the thymol contents were maximum in essential oils of T. vulgaris and O. vulgare compared to the other sample matrices studied. The thymol contents of TE of T. vulgaris and O. vulgare of different geographical regions were significantly low compared to their UBE extract. Using 12 distinct components of green analytical chemistry, the overall “analytical GREEnness (AGREE)” scale for the proposed analytical approach was computed 0.79, showing the good greener nature of the proposed analytical approach. Overall, the greener normal-phase HPTLC technique was found to be reliable for determining thymol in commercial formulations and plant-based phytopharmaceuticals.